RESUMO
Brain iron accumulation constitutes a pathognomonic indicator in several neurodegenerative disorders. Metal accumulation associated with dopaminergic neuronal death has been documented in Parkinson's disease. Through the use of in vivo and in vitro models, we demonstrated that lipid dysregulation manifests as a neuronal and glial response during iron overload. In this study, we show that cholesterol content and triacylglycerol (TAG) hydrolysis were strongly elevated in mice midbrain. Lipid cacostasis was concomitant with the loss of dopaminergic neurons, astrogliosis and elevated expression of α-synuclein. Exacerbated lipid peroxidation and markers of ferroptosis were evident in the midbrain from mice challenged with iron overload. An imbalance in the activity of lipolytic and acylation enzymes was identified, favoring neutral lipid hydrolysis, and consequently reducing TAG and cholesteryl ester levels. Notably, these observed alterations were accompanied by motor impairment in iron-treated mice. In addition, neuronal and glial cultures along with their secretomes were used to gain further insight into the mechanism underlying TAG hydrolysis and cholesterol accumulation as cellular responses to iron accumulation. We demonstrated that TAG hydrolysis in neurons is triggered by astrocyte secretomes. Moreover, we found that the ferroptosis inhibitor, ferrostatin-1, effectively prevents cholesterol accumulation both in neurons and astrocytes. Taken together, these results indicate that lipid disturbances occur in iron-overloaded mice as a consequence of iron-induced oxidative stress and depend on neuron-glia crosstalk. Our findings suggest that developing therapies aimed at restoring lipid homeostasis may lead to specific treatment for neurodegeneration associated with ferroptosis and brain iron accumulation.
Assuntos
Ferroptose , Sobrecarga de Ferro , Transtornos Motores , Camundongos , Animais , Metabolismo dos Lipídeos , Transtornos Motores/metabolismo , Ferro/metabolismo , Peroxidação de Lipídeos , Neurônios Dopaminérgicos/metabolismo , Colesterol/metabolismo , LipídeosRESUMO
BACKGROUND: 1α,25-dihydroxy vitamin D3 (calcitriol) shows potent growth-inhibitory properties on different cancer cell lines, but its hypercalcemic effects have severely hampered its therapeutic application. Therefore, it is important to develop synthetic calcitriol analogues that retain or even increase its antitumoral effects and lack hypercalcemic activity. Based on previous evidence of the potent antitumor effects of the synthetic alkynylphosphonate EM1 analogue, we have now synthesized a derivative called SG. OBJECTIVE: The aim of the present work is to evaluate the calcemic activity and the antitumor effect of SG, comparing these effects with those exerted by calcitriol and with those previously published for EM1. In addition, we propose to analyze by in silico studies, the chemical structure-biological function relationship of these molecules. METHODS: We performed the synthesis of vinylphosphonate SG analogue; in vitro assays on different cancer cell lines; in vivo assays on mice; and in silico assays applying computational molecular modeling. RESULTS: The SG compound lacks hypercalcemic activity, similar to the parent compound EM1. However, the antitumor activity was blunted, as no antiproliferative or anti-migratory effects were observed. By in silico assays, we demonstrated that SG analogue has a lower affinity for the VDRligand- binding domain than the EM1 compound due to lack of interaction with the important residues His305 and His397. CONCLUSION: These results demonstrate that the chemical modification in the lateral side chain of the SG analogue affects the antitumoral activity observed previously for EM1 but does not affect the calcemic activity. These results contribute to the rational design and synthesis of novel calcitriol analogues.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Calcitriol/química , Calcitriol/farmacologia , Organofosfonatos/química , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Desenho de Fármacos , Humanos , Relação Estrutura-AtividadeRESUMO
This Forum issue "Heme Oxygenase" (HO) includes original research articles and reviews that are aimed at understanding the role of HO-1 in several pathophysiological conditions, specially addressing those involving inflammation and oxidative damage. Overall, the seven contributions of this Forum highlight the dual role that HO-1 displays in cells and tissues, and address the molecular and cellular mechanisms through which HO-1 participates in the pathophysiology of the metabolic syndrome, obesity, cancer, and neurodegenerative, neurodevelopmental, and inflammatory bowel diseases. Indeed, one of the reviews thoroughly describes evidence of the anti-inflammatory properties of HO-1 in gut homeostasis, with potential to attenuate inflammatory bowel diseases. Three other reviews show the mostly beneficial effect of HO-1 expression in the attenuation of metabolic syndrome, obesity, cardiovascular disease, and diabetic cardiomyopathy. Contrariwise, one of the original articles show the overexpression of HO-1 in astroglia, models neurodegenerative (Parkinson-like) or neurodevelopmental (Schizophrenia-like) behaviors in mice, depending on the timing of expression of HO-1 during lifespan. The other original research communication demonstrates the role of HO-1 on the tropism of prostate cancer cells to bone, thus showing the involvement of this protein in the communication between bone and cancer cells. Finally, the Forum issue includes a review that elaborates on the classic and ultimate knowledge of HO-1 transcriptional regulation as well as the mechanisms of alternative splicing and post-transcriptional regulation of Hmox1 gene expression that have been little explored. Antioxid. Redox Signal. 32, 1239-1242.
Assuntos
Heme Oxigenase-1 , Neoplasias da Próstata , Animais , Remodelação Óssea , Heme , Heme Oxigenase (Desciclizante) , Humanos , Masculino , CamundongosRESUMO
The vitamin D receptor (VDR) constitutes a promising therapeutic target for the treatment of cancer. Unfortunately, its natural agonist calcitriol does not have clinical utility due to its potential to induce hypercalcemic effects at the concentrations required to display antitumoral activity. For this reason, the search for new calcitriol analogues with adequate therapeutic profiles has been actively pursued by the scientific community. We have previously reported the obtaining and the biological activity evaluation of new calcitriol analogues by modification of its sidechain, which exhibited relevant antiproliferative and selectivity profiles against tumoral and normal cells. In this work we conducted molecular modeling studies (i.e. molecular docking, molecular dynamics, constant pH molecular dynamics (CpHMD) and free energy of binding analysis) to elucidate at an atomistic level the molecular basis related to the potential of the new calcitriol analogues to achieve selectivity between tumoral and normal cells. Two histidine residues (His305 and His397) were found to exhibit a particular tautomeric configuration that produces the observed bioactivity. Also, different acid-based properties were observed for His305 and His307 with His305 showing an increased acidity (pKa 5.2) compared to His397 (pKa 6.8) and to the typical histidine residue. This behavior favored the pharmacodynamic interaction of the calcitriol analogues exhibiting selectivity for tumoral cells when VDR was modeled at the more acidic tumoral environment (pH â 6) compared to the case when VDR was modeled at pH 7.4 (normal cell environment). On the other hand, non-selective compounds, including calcitriol, exhibited a similar interaction pattern with VDR when the receptor was modeled at both pH conditions. The results presented constitute the first evidence on the properties of the VDR receptor in different physicochemical environments and thus represent a significant contribution to the in silico screening and design of new calcitriol analogues.
Assuntos
Modelos Moleculares , Receptores de Calcitriol/metabolismo , Calcitriol/metabolismo , Histidina/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Ligantes , Receptores de Calcitriol/química , Microambiente TumoralRESUMO
The active form of vitamin D3 , calcitriol, is a potent antiproliferative compound. However, when effective antitumor doses of calcitriol are used, hypercalcemic effects are observed, thus blocking its therapeutic application. To overcome this problem, structural analogues have been designed with the aim of retaining or even increasing the antitumor effects while decreasing its calcemic activity. This report aims at gaining insights into the structure-activity relationships of the novel oxolane-containing analogue, AM-27, recently synthesized. We herein demonstrate that this compound has antiproliferative and antimigratory effects in squamous cell carcinoma, glioblastoma, and breast cancer cell lines. Analyses of the mechanisms underlying the AM-27 effects on cell viability revealed induction of apoptosis by the analogue. Importantly, nonmalignant cell lines were little or not affected by the compound. In addition, the analogue did not produce hypercalcemia in mice. Also, in silico studies involving docking and molecular dynamics techniques showed that AM-27 is able to bind to the human vitamin D receptor with a higher affinity than the natural ligand calcitriol, a feature that is mostly derived from an electrostatic interaction pattern. Altogether, the proapoptotic effect observed in cancer cells, the lack of calcemic activity in mice, and the differential effects in normal cells suggest the potential of AM-27 as a therapeutic compound for cancer treatment.
Assuntos
Antineoplásicos/farmacologia , Calcitriol/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Calcitriol/síntese química , Calcitriol/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Simulação por Computador , Relação Dose-Resposta a Droga , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
The active form of vitamin D3, calcitriol, plays a major role in maintaining calcium/phosphate homeostasis. In addition, it is a potent antiproliferative and prodifferentiating agent. However, when effective antitumor doses of calcitriol are employed, hypercalcemic effects are observed, thus precluding its therapeutic application. To overcome this problem, structural analogues have been designed with the aim at retaining or even increasing the antitumor effects while decreasing its calcemic activity. This report shows the biological evaluation of an alkynylphosphonate vitamin D less-calcemic analogue in a murine model of breast cancer. We demonstrate that this compound has potent anti-metastatic effects through its action over cellular migration and invasion likely mediated through the up-regulation of E-cadherin expression. Based on the current in vitro and in vivo results, EM1 is a promising candidate as a therapeutic agent in breast cancer.
Assuntos
Neoplasias da Mama/patologia , Calcitriol/farmacologia , Metástase Neoplásica/prevenção & controle , Organofosfonatos/farmacologia , Animais , Calcitriol/análogos & derivados , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Vitamin D and its analogs have been shown to display anti-proliferative effects in a wide variety of cancer types including glioblastoma multiforme (GBM). These anticancer effects are mediated by its active metabolite, 1α, 25-dihydroxyvitamin D3 (calcitriol) acting mainly through vitamin D receptor (VDR) signaling. In addition to its involvement in calcitriol action, VDR has also been demonstrated to be useful as a prognostic factor for some types of cancer. However, to our knowledge, there are no studies evaluating the expression of VDR protein and its association with outcome in gliomas. Therefore, we investigated VDR expression by using immunohistochemical analysis in human glioma tissue microarrays, and analyzed the association between VDR expression and clinico-pathological parameters. We further investigated the effects of genetic and pharmacologic modulation of VDR on survival and migration of glioma cell lines. Our data demonstrate that VDR is increased in tumor tissues when compared with VDR in non-malignant brains, and that VDR expression is associated with an improved outcome in patients with GBM. We also show that both genetic and pharmacologic modulation of VDR modulates GBM cellular migration and survival and that VDR is necessary for calcitriol-mediated effects on migration. Altogether these results provide some limited evidence supporting a role for VDR in glioma progression.
Assuntos
Neoplasias Encefálicas/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Glioblastoma/metabolismo , Receptores de Calcitriol/metabolismo , Adulto , Fatores Etários , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Calcitriol/farmacologia , Agonistas dos Canais de Cálcio/farmacologia , Linhagem Celular Tumoral , Movimento Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Ciclina D1/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/mortalidade , Glioblastoma/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Oncogênicas/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Fatores Sexuais , Fatores de Tempo , Análise Serial de TecidosRESUMO
Tristetraprolin (TTP) is a RNA-binding protein that inhibits the expression of pro-inflammatory cytokines and invasiveness-associated genes. TTP levels are decreased in many different cancer types and it has been proposed that this protein could be used as a prognostic factor in breast cancer. Here, using publicly available DNA microarray datasets, "serial analysis of gene expression" libraries and qRT-PCR analysis, we determined that TTP mRNA is present in normal breast cells and its levels are significantly decreased in all breast cancer subtypes. In addition, by immunostaining, we found that TTP expression is higher in normal breast tissue and benign lesions than in infiltrating carcinomas. Among these, lower grade tumors showed increased TTP expression compared to higher grade cancers. Therefore, these data indicate that TTP protein levels would provide a better negative correlation with breast cancer invasiveness than TTP transcript levels. In mice, we found that TTP mRNA and protein expression is also diminished in mammary tumors. Interestingly, a strong positive association of TTP expression and mammary differentiation was identified in normal and tumor cells. In fact, TTP expression is highly increased during lactation, showing good correlation with various mammary differentiation factors. TTP expression was also induced in mammary HC11 cells treated with lactogenic hormones, mainly by prolactin, through Stat5A activation. The effect of this hormone was highly dependent on mammary differentiation status, as prolactin was unable to elicit a similar response in proliferating or neoplastic mammary cells. In summary, these studies show that TTP expression is strongly linked to the mammary differentiation program in human and mice, suggesting that this protein might play specific and relevant roles in the normal physiology of the gland.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Tristetraprolina/genética , Animais , Sequência de Bases , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Diferenciação Celular , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Lactação , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/crescimento & desenvolvimento , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Humanas/metabolismo , Neoplasias Mamárias Animais/genética , Neoplasias Mamárias Animais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Gravidez , Prolactina/farmacologia , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/metabolismo , Tristetraprolina/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
The expression of heme oxygenase-1 (HO-1) was shown to be increased in multiple tumors compared with their surrounding healthy tissues and was also observed to be up-regulated in oral squamous cell carcinomas (OSCC). However, conflicting results were obtained and little information is available regarding HO-1 significance in head and neck squamous cell carcinoma (HNSCC). Therefore, the aim of the present study was to perform a wide screening of HO-1 expression in a large collection of human primary HNSCCs and to correlate the results with clinical and pathological parameters. For this purpose, we investigated the expression of this protein by immunohistochemistry (IHC) in tissue microarrays (TMAs) of HNSCC and in an independent cohort of paraffin-embedded tumor specimens. HO-1 expression was further validated by real-time qPCR performed on selected laser capture-microdissected (LCM) oral tissue samples. Both the number of HO-1-positive samples and HO-1 immunoreactivity in the cancerous tissues were significantly higher than those in the non-tumor tissues. These results were confirmed at the mRNA level. Interestingly, HO-1 localization was observed in the nucleus, and the rate of nuclear HO-1 in HNSCC was higher than that in non-malignant tissues. Nuclear HO-1 was observed in HNSCC cell lines and increased even further following hemin treatment. Analysis of HO-1 expression and sub-cellular localization in a mouse model of squamous cell carcinoma (SCC) and in human HNSCC revealed that nuclear HO-1 increases with tumor progression. Taken together, these results demonstrate that HO-1 is up-regulated in HNSCC and that nuclear localization of HO-1 is associated with malignant progression in this tumor type.
Assuntos
Carcinoma de Células Escamosas/enzimologia , Neoplasias de Cabeça e Pescoço/enzimologia , Heme Oxigenase-1/metabolismo , Idoso , Animais , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/genética , Modelos Animais de Doenças , Progressão da Doença , Feminino , Neoplasias de Cabeça e Pescoço/diagnóstico , Neoplasias de Cabeça e Pescoço/genética , Heme Oxigenase-1/genética , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Inclusão em Parafina , Reação em Cadeia da Polimerase , Prognóstico , RNA Mensageiro/metabolismo , Análise Serial de TecidosRESUMO
While changes in heme oxygenase (HO-1) in lung cancer have already been reported, conflicting results were obtained for enzyme expression in human lung cancer specimens. Therefore, the aim of this work was to study HO-1 expression in a large collection of human lung cancer samples. For this purpose, we analyzed the expression of HO-1 in an organized tissue microarray (TMA) and investigated its correlation with clinicopathological data. Ninety-six percent of tumor samples were positive for HO-1, and the expression of HO-1 was significantly higher in cancerous than in non-cancerous tissues. Importantly, HO-1 expression correlated with advanced stages and lymph node involvement. Additionally, quantitative RT-PCR in 18 pairs of human lung carcinomas and their adjacent non-malignant tissues was performed. Our results demonstrate that HO-1 protein is upregulated in epithelial malignant cells in NSCLC and its expression is associated with higher stages of the disease. Additionally, different subcellular localization is observed between tumor and adjacent non-malignant tissues.